Objective:To analyze the effects of espO gene knockout on the biological characteristics of enterhemorrhagic Escherichia coli (EHEC). Methods:Two-step methods mediated by the suicide plasmid pCVD442-Δ espO and plasmid pTrc99a were used to construct the espO gene-deleted strain (Δ espO) and the complemented mutant (CΔ espO), respectively. HeLa cells were infected with different EHEC strains to analyze the biological functions and lethal effects of espO gene during infection. Results:PCR, electrophoresis and gene sequencing showed that the Δ espO and CΔ espO mutants were successfully constructed. Compared with the wild-type strain, neither the Δ espO nor CΔ espO mutant showed significant difference in growth rate, indicating that the espO gene had no influence on the growth and replication of EHEC. Furthermore, EspO could activate the tumor necrosis factor receptor (TNF)-induced NF-κB signaling pathway, while the effector protein NleB could inhibit the process. EspO could not inhibit the death of HeLa cells induced by TNF or TNF-related apoptosis-inducing ligand (TRAIL) after EHEC infection. Conclusions:In this study, we successfully constructed the espO gene-deleted and complemented mutants of EHEC and preliminarily analyzed the interaction between espO gene and host cells and the effects of espO gene on cell apoptosis during infection, which provided reference for further research on the in vitro biochemical activity and in vivo pathogenic roles of EspO.

Objective To investigate the application of venereal disease research laboratory(VDRL) test and toluidine red unheated serum test(TRUST) in syphilis laboratory diagnosis.Methods Serum,plasma and cerebrospinal fluid (CSF) in syphilis patients and healthy controls were measured by VDRL and TRUST.Results The VDRL detection results in serum and CSF sepcimens were generally higher than the TRUST detection results by 1-2 titers,while in plasma specimen,the VDRL detection results were generally lower than the TRUST detection results by 1-2 titers than TRUST when using plasma specimen.In addition,the VDRL detection in normal control serum and plasma specimens all appeared different degrees of false positive,but in the detection of normal control CSF,the results of TRUST and VDRL were consistent.Conclusion TRUST is more suitable for serum and plasma specimens,and CSF is suitable for the CSF specimen,but not suitable for serum and plasma detection.

Objective To investigate the expression of PD-1 on CD4 +and CD8 + T cells in patients with HBeAg-positive chronic hepatitis B(CHB)treated with telbivudine.Methods Fifty-six HBeAg-positive CHB patients admitted in the First Affiliated Hospital of Zhengzhou University during January 201 3 and June 201 4 were enrolled in this study.The expression of PD-1 on CD4 +and CD8 + T cells was detected with flow cytometry at baseline,24,48 and 72 wks after telbivudine treatment.The relationship of PD-1 expression with alanine aminotransferase (ALT)level,HBeAg seroconversion and HBV DNA loads was analyzed.t test and completely random variance analysis were used to analyze the data.Results The PD-1 expression on CD4 + and CD8 + T cells at baseline was higher in patients with low ALT levels compared to those with high ALT levels(t =1 2.20 and 9.69,both P <0.01 ),while higher levels of PD-1 expression was also observed in patients with high HBV DNA load (≥5 lgIU /mL)compared to those with low HBV DNA load (t =4.39 and 4.85,both P <0.01 ).PD-1 levels on CD4 + and CD8 + T cells presented a declining trend after telbivudine treatment(F =6.98 and 8.97,both P <0.01 ),PD-1 expression in patients with HBeAg seroconversion showed lower levels compared with baseline values (t =1 8.45 and 1 8.01 ,both P <0.01 ). Conclusion In HBeAg-positive CHB patients,the expression of PD-1 on CD4 + and CD8 + T lymphocytes shows a decreasing trend during the treatment with telbivudine,indicating that antiviral therapy may alleviate the immunosuppression in these patients.

BACKGROUND: Topoisomerase II alpha (TOP2A) has been reported to play a crucial role in the tumorigenesis of various cancer types. However, the biological role of TOP2A in gallbladder cancer (GBC) remains unknown. The current study aimed to explore the function and potential mechanism of TOP2A in GBC. METHODS: Based on Gene Expression Profiling Interactive Analysis data, we found TOP2A was significantly up-regulated in GBC tissues and resulting in shorter overall survival. Quantitative real-time polymerase chain reaction and immunohistochemistry were conducted to detect the expression of TOP2A in 45 pairs of GBC tissues and adjacent non-tumor tissues. In vitro, cell proliferation, migration, and invasion ability were examined by cell counting kit-8 and transwell assay, respectively. Epithelial-mesenchymal transition (EMT) related and phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/Akt/mTOR) pathway-related markers were measured by Western blotting. Xenograft model assay was performed to evaluate the effect of TOP2A in vivo. RESULTS: TOP2A was found up-regulated in GBC (tumor vs. normal, 12.62 vs. 0.34) and correlated with the late tumor node metastasis stage (P = 0.0032), present of lymph node metastasis (P = 0.0273), and poor prognosis in GBC patients (log-rank P = 0.028). In vitro and in vivo assays showed that knockdown of TOP2A notably inhibited cell proliferation, migration, invasion, EMT process, and tumor growth in GBC. In addition, TOP2A down-regulation significantly decreased the protein levels of phosphor (p)-PI3K, p-Akt, and p-mTOR. CONCLUSION Our study demonstrates that TOP2A was overexpressed in GBC and associated with poor prognosis in GBC patients. TOP2A promotes GBC cell proliferation, migration, invasion, EMT process, and tumor growth through activating PI3K/Akt/mTOR signaling pathway, and may serve as a novel prognostic biomarker and therapeutic target for GBC.

Objective To explore current status and influencing factors of nurse career attitude in one primary hospital in Chongqing. Methods By nurse career attitude scale made by Nursing School of the Second Military Medical University, a total of 213 nurses had been chosen and investigated by stratified sampling method from our hospital. The result of investigation had been analyzed by descriptive statistics analysis method, single factor analysis of variance and multiple liner regression analysis. Results The total career attitude score of nurse in primary hospital acquired 2. 01 ~ 8. 36, average score was (4. 33 ± 0. 88) . In thirty clauses, the most people chose long-term three shifts work arranged by manager, harmful personal health education, impacting on family life occupying 75. 59% (161 / 213). By single factor analysis of variance, it showed different level of career attitude that nurses were in different gender, educational level, occupational title, income level and three shifts (F = 11. 35,8. 30,3. 16,8. 44,3. 53, respectively; P < 0. 05). The result of multiple liner regression analysis indicated that nurses had more positive attitudes with male, high education, high occupational title, high income, less three shifts comparing with others (P < 0. 05). Conclusions Both internal and external causes working together formed positive career attitudes of nurses in primary hospital. The administrative department should establish a harmonious environment, guide nurses correctly, and inspire them by career education.

Objective The study was carried out to investigate the effects of different dietary Ca levels on bone metabolism, based on the osteoprotegerin ( OPG)-receptor activator of NF?κB ligand ( RANKL)?receptor activator of NF?κB ( RANK) system in growing WHBE rabbits. Methods Twenty one weaned male WHBE rabbits at the age of 42 days were divided into 3 groups (I, II and III) according to dietary Ca levels (0. 95%, 1. 10%, and 1. 30%, respectively) for a 42?d feeding trial. The above three diets had similar phosphor (P) (about 0. 64 g/kg), digestible energy (9. 50 MJ/kg), crude protein (about 19. 70%) and crude fibre (13. 57%) contents. When the feeding trial finished, the serum in?dices of bone metabolism (Ca, PTH and BALP) were detected, and the OPG?RANK?RANKL system in bone tissue was analyzed by real?time fluorescence quantitative PCR assay and immunohistochemistry, respectively. At last, the relationships between bone metabolism and dietary Ca were evaluated according to RANKL/OPG ratio. Results All the contents of serum Ca, PTH and BALP had no significant differences in the groups I, II and III (P>0. 05). Both the RANKL mR?NA and RANKL/OPG mRNA ratio were lowest in the group II and had significant differences with group I and III ( P<0. 05). Dietary Ca levels had significant effects on the protein expression of OPG?RANK?RANKL in bone tissues (P<0. 01). The positive index of OPG in the groups II and III was significantly higher than that in the group I(P<0. 01), while the positive index of RANK in the group II was lower than those of the group I and III(P<0. 01). The protein ex?pression positive index ratio of RANKL to OPG was also lowest in the group II, showing a significant difference with group I(P<0. 01). Furthermore, both the gene transcription ratio of RANKL to OPG and the protein expression positive index ratio of RANKL to OPG had significant correlations (with quadratic curve) to the dietary Ca levels (R2 =0. 4068, 0. 8433;P<0. 05,P<0. 001). Conclusions In summary, the bone metabolism of WHBE rabbits during growing periods has sig?nificant correlation with dietary Ca levels. An optimal bone metabolism status can be obtain at 1. 10% dietary Ca level as demonstrated in this study.

Objective To evaluate the efficacy of quadrates lumborum block for unilateral inguinal hernia repair in elderly patients. Methods Fifty-eight elderly patients with unilateral inguinal hernia of both sexes, aged 65-80 yr, with body mass index of 18-25 kg∕m2 , of American Society of Anesthesiolo-gists physical status Ⅱ or Ⅲ, scheduled for elective unilateral tension-free repair, were divided into 2 groups ( n=29 each) using a random number table method: iliohypogastric-ilioinguinal nerve block group (group T) and quadrates lumborum block group (group Q). Iliohypogastric-ilioinguinal nerve block with arteria circumflexa ilium profunda as a marker was carried out with 0. 33％ ropivacaine 20 ml under ultra-sound guidance in group T. The anterior approach to quadratus lumborum block was performed with 0. 33％ropivacaine 20 ml under ultrasound guidance in group Q. Operation was started after the height of sensory block was assessed by pin-prick test at 30 min after block. When the blocking effect did not meet the opera-tion requirements, an increment of 1％ lidocaine 2. 5 ml was given every time in the surgical field until op-eration requirements were met. Dexmedetomidine was intravenously infused at a rate of 0. 03-0. 07μg·kg-1 ·min-1 during surgery until the end of surgery to maintain Narcotrend index between 80 and 90. When postoperative visual analogue scale score >3, parecoxib sodium 40 mg was intravenously injected, and if marked pain relief was not found 10 min later, tramadol hydrochloride 50-100 mg was intravenously injected. The upper spread of sensory block and intraoperative requirement for additional local anesthetics were recorded at 30 min after nerve block. The requirement for parecoxib and tramadol was recorded within 48 h after operation. The development of inadvertent intravascular injection of local anesthetics, local anes-thetic intoxication and postoperative nausea and vomiting, nerve block of lower extremity and uroschesis was recorded. Results Skin pain disappeared at the plane of T11-L1 in group T and at the plane of T9-L1 in group Q. Compared with group T, the intraoperative requirement for and consumption of local anesthetics, postoperative requirement for parecoxib and tramadol, and postoperative incidence of nausea and vomiting were significantly decreased in group Q ( P<0. 05) . Conclusion Quadrates lumborum block provides bet-ter efficacy for unilateral inguinal hernia repair than iliohypogastric-ilioinguinal nerve block in elderly pa-tients.

Background:Blood glucose control is closely related to type 2 diabetes mellitus (T2DM) prognosis.This multicenter study aimed to investigate blood glucose control among patients with insulin-treated T2DM in North China and explore the application value of combining an elastic network (EN) with a machine-learning algorithm to predict glycemic control.Methods:Basic information,biochemical indices,and diabetes-related data were collected via questionnaire from 2787 consecutive participants recruited from 27 centers in six cities between January 2016 and December 2017.An EN regression was used to address variable collinearity.Then,three common machine learning algorithms (random forest [RF],support vector machine [SVM],and back propagation artificial neural network [BP-ANN]) were used to simulate and predict blood glucose status.Additionally,a stepwise logistic regression was performed to compare the machine learning models.Results:The well-controlled blood glucose rate was 45.82％ in North China.The multivariable analysis found that hypertension history,atherosclerotic cardiovascular disease history,exercise,and total cholesterol were protective factors in glycosylated hemoglobin (HbA1c) control,while central adiposity,family history,T2DM duration,complications,insulin dose,blood pressure,and hypertension were risk factors for elevated HbA1c.Before the dimensional reduction in the EN,the areas under the curve of RF,SVM,and BP were 0.73,0.61,and 0.70,respectively,while these figures increased to 0.75,0.72,and 0.72,respectively,after dimensional reduction.Moreover,the EN and machine learning models had higher sensitivity and accuracy than the logistic regression models (the sensitivity and accuracy of logistic were 0.52 and 0.56;RF:0.79,0.70;SVM:0.84,0.73;BP-ANN:0.78,0.73,respectively).Conclusions:More than half of T2DM patients in North China had poor glycemic control and were at a higher risk of developing diabetic complications.The EN and machine learning algorithms are alternative choices,in addition to the traditional logistic model,for building predictive models of blood glucose control in patents with T2DM.

Objective To evaluate the efficacy and safety of epalrestat, an aldose reductase inhibitor, and epalrestat plus methylcobalamine on diabetic peripheral neuropathy, as compared with methylcobalamine. Methods A total of 444 subjects with diabetic neuropathy were enrolled in the study, and divided into methylcobalamine group ( n= 145 ) , epalrestat group ( n = 143 ) , and methylcobalamine combined with epalrestat group ( n = 156 ) . Therapeutic efficacay was assessed in terms of clinical symptoms and physical examinations by using Michigan Neuropathy Screening Instrument ( MNSI ) , and electrophysiological assessments. Results After 4 to 12-weeks′treatment, symptoms and signs of neuropathy ( using MNSI ) are significantly improved in the three groups ( P<0. 01). The mean changes of MNSI ( questionnaire) score from baseline were higher in epalrestat group and methylcobalamine combined with epalrestat group as compared with that of methylcobalamine group(P<0. 05), but no difference was detected in the change of MNSI ( physical examination ) score from baseline among three groups. After treatment for 12 weeks, motor nerve conduction velocity ( MNCV ) was significantly improved in epalrestat group and methylcobalamine combined with epalrestat group(P<0. 05), but no difference was detected in MNCV at 12 week among three groups(P>0. 05). Conclusion Epalrestat is effective and safe in the treatment of diabetic neuropathy. Furthermore, epalrestat is more efficacious in ameliorating symptoms and MNCV of neuropathy than methylcobalamine. However, while no improved efficacy is shown with the combined treatment.

Objective To isolate and culture a clinical strain (GDI) of Treponema pallidum (Tp) in Guangdong province,and to investigate the difference in nonsynonymous single nucleotide polymorphisms (nsSNPs) between the GD1 strain and Tp Nichols strain.Methods The GD1 strain was isolated from the hard chancre in a patient with primary syphilis in Guangdong province,and continuously subcultured in the testes of New Zealand white rabbit.The serial subcultivation of GD1 was multi-verified by dark-field microscopy,polymerase chain reaction (PCR) for TP0548 gene,DNA sequencing and genotyping.Meglumine diatrizoate density gradient centrifugation was performed to isolate rabbit tissues and concentrate GD1,and DNA sequencing was used to verify the nsSNPs in the TP0443 and TP0584 genes.Results The GD1 strain was successfully isolated from the lesions of the patient with syphilis,and classified as a subtype f of TP0548.Compared with the American Tp (Nichols strain),there were nsSNP mutations in the GD1 strain.One mutation was located in the TP0443 gene,leading to the the substitution of threonine by alanine at amino acid position 120,and another one was located in the TP0584 gene,which caused a change from alanine to threonine at amino acid position 314.Conclusion The GD1 strain was successfully isolated firstly from the lesions in a patient with syphilis in Guangdong province,and nsSNP mutations were confirmed in the GD 1 strain on the etiology.